We prepare a wide variety of nucleotide analogs for protein-binding studies; their chemical and spectral properties are described in Probes for Protein Kinases, Protein Phosphatases and Nucleotide-Binding Proteins—Section 17.3. These include various fluorescent, photoaffinity and caged versions of adenosine and guanosine triphosphates, diphosphates and cyclic monophosphates. The GTP analogs are among the most important probes for the study of G proteins and G protein–coupled receptors (GPCR). Heterotrimeric guanine nucleotide–binding regulatory proteins transmit a variety of receptor signals to modulate diverse cellular responses, including apoptosis.

G proteins are composed of α-, β- and γ-subunits. Upon receptor stimulus, the α-subunit of the heterotrimeric G proteins exchanges GDP for GTP and dissociates from the β-γ-subunit complex. The GTP-bound G protein will interact with various second messenger systems, either inhibiting (Gi) or stimulating (Gs) their activity. Stimulatory G proteins are permanently activated by cholera toxin, inhibitory G proteins by pertussis toxin. The α-subunit has a slow intrinsic rate of GTP hydrolysis, and once the GTP is hydrolyzed it reassociates with the β-γ-subunit complex. The GTP hydrolysis by G proteins is regulated by interactions with GTPase-activating proteins, or GAPs. There is a large family of GAPs for G proteins known as regulators of G protein signaling or RGS proteins. G proteins are turned off when the α-subunit hydrolyzes the GTP, either spontaneously or upon interaction with a GTPase-activating protein, permitting the heterotrimeric α-β-γ-complex to reassociate.

The GAPs are a diverse group of monomeric GTPases, including ARF, Ran, Ras, Rab, Rac, Rho and Sar, which play an important part in regulating many intracellular processes, such as cytoskeletal organization and secretion. There is less diversity among the β- and γ-subunits, but they may have direct activating effects in their own right. Most β- and γ-subunits are post-translationally modified by myristoylation or isoprenylation, which may alter their association with membranes.